Engineering Molecular Motors Molecular motors lie at the heart of biological processes from DNA replication to cell migration. The principal goal of my research is to understand the physical mechanisms by which these nanoscale machines convert chemical energy into mechanical work. I propose a radical change in the way my laboratory approaches this goal. We will rigorously challenge our understanding of the relationships between molecular structures and mechanical functions by rationally engineering molecular motors with novel properties. The performance of our designs will illuminate both the inner workings of natural biological motors and the general operational constraints for producing directed motion on the molecular scale. Ultimately, construction of molecular motors to arbitrary specifications will provide a powerful toolkit for synthetic biology, therapeutics, and nanotechnology. My laboratory will design and characterize molecular motor variants using a rapid testing cycle that relies on new instrumentation for high throughput single molecule tracking and manipulation assays. We will focus our efforts by choosing a small number of ambitious design targets, each requiring several intermediate molecular innovations and optimizations. For the period of this award, two initial design targets will be pursued, leveraging our existing expertise in myosin and topoisomerase mechanochemistry: 1) We will create a fast, processive myosin motor that can be optically or chemically signaled to reversibly switch its direction of hand-over-hand movement along an actin filament. 2) We will create a robust tension-insensitive rotary DNA-associated motor that introduces torque in the opposite direction from DNA gyrase. Success will represent an unprecedented level of control over nanoscale motion, building an engineering capacity that will eventually be used to design protein nanoassemblies capable of sophisticated intracellular therapeutic functions such as genome repair. Novel molecular motors will also have ex vivo applications including molecular sorting and assembly of nanoelectronics in microfabricated devices.